14 Lecture Animation Ppt

1. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. DNA probe DNA probe array tagged DNA tagged DNA did bind to probe tagged DNA did not bind to probe testing subject's DNA

2. Outline DNA Cloning Recombinant DNA Technology Restriction Enzyme DNA Ligase Polymerase Chain Reaction Biotechnology Products Transgenic Bacteria, Plants, and Animals Gene Therapy Genomics

3. DNA Cloning Cloning is the production of identical copies of DNA Members of a bacterial colony on a petri dish are clones because they all came from division of the same cell. Identical twins are clones Single embryo separate to become two. Gene cloning is production of many identical copies of the same gene. If the inserted gene is replicated and expressed, we can recover the cloned gene or protein product. Cloned genes have many research purposes Humans can be treated with gene therapy

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5. Recombinant DNA Technology Recombinant DNA (rDNA) contains DNA from two or more different sources Requires: A vector introduces rDNA into host cell Plasmids (small accessory rings of DNA from bacteria) are common vectors Two enzymes are required to introduce foreign DNA into vector DNA A restriction enzyme - cleaves DNA, and A DNA ligase enzyme - seals DNA into an opening created by the restriction enzyme

7. Cloning a Human Gene Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. O f r o m a i 1. Restriction enzyme cleaves DNA. 2. DNA ligase seals human gene and plasmid. 3. Host cell takes up recombined plasmid. 4. Gene cloning occurs. bacterium recombinant DNA insulin human cell insulin gene human DNA plasmid

8. Restriction Enzyme Cuts DNA at specific points. Cleaves vector (plasmid) and foreign (human) DNA. Cleaving DNA makes DNA fragments ending in short single-stranded segments with “sticky ends.” The “sticky ends” allow insertion of foreign DNA into vector DNA. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. DNA duplex "sticky ends" restriction enzyme A T A T A T A T A T C G C C G G C G A T A T A T A T A T C G C C G G C G

10. DNA Ligase Seals the foreign gene into the vector DNA Treated cells (bacteria) take up plasmids Bacteria and plasmids reproduce. Many copies of the plasmid and many copies of the foreign gene.

14. DNA Cloning: Polymerase Chain Reaction (PCR) Amplifies a targeted sequence of DNA Create millions of copies of a single gene or a specific piece of DNA in a test tube Requires: DNA polymerase Withstands the temperature necessary to separate double-stranded DNA. A supply of nucleotides for the new, complementary strand

15. PCR Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. PCR cycles DNA copies first 1 second 2 third 4 old old old strand new new new strand DNA double strand fourth 8 fifth 16 and so forth

19. Applications of PCR: Analyzing DNA Segments DNA fingerprinting is the technique of using DNA fragment lengths Treat DNA segment with restriction enzymes A unique collection of different fragments is produced Gel electrophoresis separates the fragments according to their charge/size Produces distinctive banding pattern Usually used to measure number of repeats of short sequences Used in paternity suits, rape cases, corpse ID, etc.

21. DNA Fingerprinting & Paternity Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. Mother Child Male 1 Male 2 Base repeat units many few DNA Band patterns Increasing size b. Automated DNA fingerprinting a. Fluorescence units

23. Biotechnology Products Genetically engineered organisms can produce biotechnology products. Organisms that have had a foreign gene inserted into them are transgenic .

25. Transgenic Bacteria Gene of interest is inserted into bacteria. Bacteria are grown in large vats called bioreactors and product is harvested. Products on the market include insulin, hepatitis B vaccine, t-PA, and human growth hormone. Transgenic bacteria can produce chemical products. Transgenic bacteria process minerals. Metals Collection

26. Transgenic Bacteria Transgenic bacteria promote plant health Bacteria that colonize corn roots can be endowed with genes for insect toxin Transgenic bacteria can degrade substances. Oil-Eating Bacteria

29. Transgenic Plants Agricultural Crops Foreign genes now give cotton, corn, and potato strains the ability to produce an insect toxin Soybeans are now resistant to a common herbicide Human Hormones Plants are being engineered to produce human proteins including hormones, clotting factors, and antibodies in their seeds

31. Transgenic Animals Vortex Mixing: Many types of animal eggs have taken up the gene for bovine growth hormone (bGH) The procedure has been used to produce larger fishes, cows, pigs, rabbits, and sheep Gene Pharming: Use of transgenic farm animals to produce pharmaceuticals Genes coding for therapeutic & diagnostic proteins are incorporated into an animal’s DNA The proteins appear in the animal’s milk Plans are to produce drugs to treat Cystic fibrosis Cancer Blood diseases, etc.

32. Transgenic Mammals Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. human gene for growth hormone human growth hormone microinjection of human gene development within a host goat Transgenic goat produces human growth hormone. a. b. transgenic goat cells with gene for human growth hormone microinjection of transgenic gene into enucleated donor eggs donor of eggs donor of egg enucleated eggs Cloned transgenic goats produce human growth hormone. milk development within host goats milk

33. Transgenic Animals Researchers are using transgenic mice for various research projects. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. one-celled mouse embryos with two X chromosomes Embryo develops into a female. FEMALE Embryo develops into a male. MALE no injection inject SRY DNA

34. Gene Therapy Gene therapy involves procedures to give patients healthy genes to make up for a faulty gene. It also includes the use of genes to treat genetic disorders and various human illnesses. There are ex vivo (outside body) and in vivo (inside body) methods of gene therapy. Ex Vivo Children with Severe Combined Immunodeficiency Bone Marrow Stem Cells In Vivo Cystic Fibrosis Nasal / Respiratory Spray

35. Gene Therapy Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. Brain (gene transfer by injection)* • Huntington disease • Alzheimer disease • Parkinson disease • brain tumors Skin (gene transfer by modified blood cells)** • skin cancer Lungs (gene transfer by aerosol spray)* • cystic fibrosis • hereditary emphysema Liver (gene transfer by modified implants)** • familial hypercholesterolemia Blood (gene transfer by bone marrow transplant)* • sickle-cell disease Muscle (gene transfer by injection)* • Duchenne muscular dystrophy Bone marrow (gene transfer by implantation of modified stem cells)** • SCID • sickle-cell disease Endothelium (blood vessel lining) (gene transfer by implantation of modified implants)** • hemophilia • diabetes mellitus * in vivo ** ex vivo

36. Genomics Genomics is the study of genomes of humans and other organisms. Sequencing the Bases The Human Genome Project produced a working draft of all the base pairs in all chromosomes. Took 13 years to sequence three billion base pairs along the length of chromosomes.

37. Human Genome Project Genome - All the genetic information of an individual (or species) Goals of Human Genome Project Determine the base pair sequence Construct a map showing sequence of genes on specific chromosomes

38. Human Genome Project Humans have 20,500 genes Most code for proteins Much of the human genome was formerly described as “junk” Does not specify the order of amino acids in a polypeptide Recent observation suggest that between 74% and 93% of the genome is transcribed into RNA Thus, vast junk DNA wasteland may be much more important than once thought

39. Eukaryotic Gene Structure Historically, genes were defined as discrete units of heredity that corresponded to a locus on a chromosome. Prokaryotes typically possess a single circular chromosome. Eukaryotic chromosomes are much more complex.

40. Chromosomal DNA Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. exon exon intron intron exon exon intergenic sequences DNA pre-mRNA RNA introns Gene A Gene B Gene C Gene A mRNA Gene B mRNA Gene C mRNA © Cindy Charles/PhotoEdit 5  3  5  3  3  5 

41. Eukaryotic Gene Structure An Intergenic Sequences are DNA sequences that occur between genes Repetitive DNA elements occur when the same sequence of two or more nucleotides are repeated many times along the length of one or more chromosomes. Transposons are specific DNA sequences that have the remarkable ability to move within and between chromosomes .

46. HapMap Project People inherit patterns of sequence differences, called haplotypes If one haplotype of a person has an A rather than a G at a particular location in a chromosome, there are probably other particular base differences near the A Genetic data from African, Asian, and European populations will be analyzed A HapMap is a catalog common sequence differences that occur in a species The goal of the project is to link haplotypes to risk for specific illnesses May lead to new methods of preventing, diagnosing, and treating disease

47. Functional and Comparative Genomics Functional genomics aims to understand the role of genome in cells or organisms DNA microarrays can monitor the expression of thousands of genes simultaneously and tell us: What genes are turned on Environmental conditions that turn on the gene DNA microarrays contain microscopic amounts of known DNA fixed onto a small glass slide mRNA bind through complementary base pairing mRNA is produced by active cell. Identify various mutations in the genome of an individual This is called the person’s genetic profile.

48. DNA Microarray Technology Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. DNA probe DNA probe array tagged DNA tagged DNA did bind to probe tagged DNA did not bind to probe testing subject's DNA

50. Genetic Profile The complete genotype of an individual This is the person’s genetic profile A way of studying how genes work together to control the phenotype Analyze the genetic profile of many individuals Compare their profiles to their phenotypes

51. Proteomics The study of the structure, function, and interaction of cellular proteins At least 25,000 of our genes are translated into proteins The sum total of these proteins is called the human proteome Understanding protein function is essential to the development of better drugs Correlate drug treatment to the particular genome Increase efficiency and decrease side effects Once the primary structure of these protein is known It should be possible to predict their tertiary structure Computer modeling of the tertiary of these proteins is an important part of proteomics

52. Bioinformatics The application of computer technologies to the study of the genome Genomics and proteomics produce raw data These fields depend on computer analysis to find significant patterns in the data Scientists hope to find relationships between genetic profiles and genetic disorders New computational tools will be needed to accomplish these goals

53. Bioinformatics Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. exon exon intron intron exon exon intergenic sequences DNA pre-mRNA RNA introns Gene A Gene B Gene C Gene A mRNA Gene B mRNA Gene C mRNA 5  3  5  3  3  5 

54. Review DNA Cloning Recombinant DNA Technology Restriction Enzyme DNA Ligase Polymerase Chain Reaction Biotechnology Products Gene Therapy Genomics

55. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. DNA probe DNA probe array tagged DNA tagged DNA did bind to probe tagged DNA did not bind to probe testing subject's DNA

14 Lecture Animation Ppt

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